The sense of taste can be divided into five predominant categories: bitter, salty, sour, sweet and umami. Taste perception begins with the interaction of sapid molecules (tastants) with taste receptor cells, which reside in specialized structures known as taste buds (Margolskee et al., 1993, Bioessays 15:645-50; Margolskee et al., 1993, Current Opin Neurobiol. 3:526-31). Taste buds are located in the papillae of the tongue and are the end organ of taste. The function of the taste buds is to relay information on the nutrient content of food to the central nervous system through afferent gustatory fibers. Recent advances in the molecular biology and biochemistry of taste have shown that each taste modality affects receptor cells through distinct mechanisms (Gilbertson et al., 2000 Curr Opin Neurobiol 10:519-27). Transduction of responses to bitter and sweet compounds are predominantly mediated via guanine nucleotide binding protein (G protein) coupled receptors, while salty and sour responses involve interactions with ion channels.
Gustducin is a taste selective G protein (McLaughlin et al., 1992 Nature 357:563-9). Gustducin is homologous (˜80% identical/˜90% similar) to transducin, the G protein of the visual system, and both gustducin and transducin have been immunocytochemically localized to taste buds. In the retina, light activates rhodopsin, a member of the seven transmembrane-helical G protein coupled receptor family, resulting in a conformational change and activation of transducin. Transducin subsequently disinhibits a cyclic guanidine monophosphate (cGMP) specific phosphodiesterase (PDE) and the resultant decreased cGMP concentration leads to modulation of ion channel permeability causing rod cell hyperpolarization (Birnbaumer et al, 1990 Biochem, Biophys. Acta 1031:163-224). Because of gustducin's high degree of similarity to transducin, it is thought that gustducin may be involved in taste signal transduction by modulation of a taste specific PDE. This hypothesis is further supported by the fact that transducin's PDE activating domain is 86% identical and 95% similar to gustducin's, while other G proteins have much lower relatedness in this region (Rarick et al., 1992 Science 256:1031-3). Furthermore, recombinant gustducin expressed in SF9 cells has been shown to be activated by rhodopsin and can activate retinal and taste cGMP PDE (Hoon et al., 1995 Biochem J 309:629-36; Ruiz-Avila et al., 1995 Nature 376:80-5). Thus, gustducin and transducin appear to be interchangeable in this regard.
Transducin has also been immunocytochemically localized to taste buds, and has been implicated in taste signal transduction by activation of a taste specific PDE activity (Ruiz-Avila et al., 1995 Nature 376:80-5). Furthermore, this study and subsequent work (Ming et al., 1998 Proc Nat'l Acad Sci USA 95:8933-8) demonstrated that taste bud containing membranes from bovine circumvallate papillae activated exogenously added transducin in response to bitter stimuli including denatonium, quinine, strychnine, atropine and naringen.
Gustducin has been implicated in vivo in transducing responses to bitter and sweet compounds (Wong et al., 1996, Nature 381:796-800). Gene replacement was used to generate a null mutation of the α-gustducin gene in mice. The α-gustducin knockout mice were shown to be deficient in responses to both bitter and sweet compounds as measured by two bottle preference tests as well as electrophysiology. The gustducin knockout had no effect on responses to sour or salty compounds.
Recently, putative human and rodent taste receptors for bitter taste (the T2Rs) have been cloned and cells expressing certain of these clones demonstrated to respond to the bitter compounds denatonium, cyclohexamide and 6-n-propyl-2-thiouracil (PROP) (Hoon et al., 1995 Biochem J 309:629-36; Adler et al., 2000 Cell 100:693-702; Chandrashekar et al., 2000 Cell 100:703-11; Matsunami et al., 2000 Nature 404:601-4). The T2R receptors appear to be specifically expressed in only the α-gustducin positive taste receptor cells, consistent with their proposed role in bitter transduction.
Although gustducin- and transducin-mediated pathways appear to be the primary mechanism by which responses to bitter compounds are transduced, alternative mechanisms have also been proposed. Evidence thus far suggests that bitter taste transduction may be mediated by (1) G protein coupled receptors acting via gustducin/transducin (Ruiz-Avila et al., 1995 Nature 376:80-5; Wong et al, 1996; Ming et al., 1998 Proc Nat'l Acad Sci USA 95:8933-8; Gravina et al., 2001, in preparation). Our work and that of others suggest that at least 50% of bitter compounds couple through a receptor-dependent gustducin/transducin pathway; (2) G protein coupled receptors acting via Gq or βγ subunits to generate inositol triphosphate (Spielman et al., 1994 Physiol Behav 56:1149-55; Huang et al., 1999 Nat Neurosci 2:1055-62). A recently identified G protein γ subunit expressed in gustducin positive taste cells has been shown to mediate the response of certain bitter compounds to a phospholipase C (PLC) catalyzed increase in inositol triphosphate (IP3) (Huang et al., 1999 Nat Neurosci 2:1055-62). This γ subunit is associated with gustducin in the taste cell. Other tastants appear to link via a different G protein (Gq) to IP3 production (Spielman et al., 1994 Physiol Behav 56:1149-55); (3) Receptor independent effects of bitter tasting molecules acting directly on G proteins and effector proteins such as phosphodiesterase and ion channels (Naim et al., 1994 Biochem J 297:451-4; Amer and Kreighbaum, 1975 J. Pharm. Sci. 64:1-35; Tsunenari, 1999 J Physiol 519 Pt 2:397-404).
Traditionally, sweeteners and flavorants have been used to mask the bitter taste of pharmaceuticals. The sweetener or flavorant are known to activate other taste pathways and at sufficiently high concentration this serves to mask the bitter taste of the pharmaceutical. However, this approach has proved ineffective at masking the taste of very bitter compounds. Microencapsulation in a cellulose derivative has also been used to mask the bitter taste of pharmaceuticals, however this approach prevents rapid oral absorption of the pharmaceutical. The nucleotide monophosphates IMP and GMP have been used to counter the metallic or pseudo-bitter taste of KCl for its use in low sodium edible salt composition (Zolotov et al., U.S. Pat. No. 5,853,792). In addition, AMP has been described as an inhibitor of bitter taste (Ming et al., Proc. Natl Acad Sci USA 96:9903-8; McGregor and Gravina 2001, 23rd Meeting of the Association of Chemoreception Sciences).